Proposal of a molecular testing algorithm for differentiated thyroid cancer (DTC).
Authors: Margherita Nannini, Andrea Repaci, Antonio De Leo, Valentina Vicennati, Elisa Gruppioni, Annalisa Altimari, Erica Solaroli, Elisa Lodi Rizzini, Fabio Monari, Gabriella Savastio, Luigi Lovato, Giovanni Tallini, Maria A. Pantaleo, Dario De Biase
Journal: Journal of Clinical Oncology (2022)

e18090 Background: In the growing era of precision medicine and tumor-agnostic therapies, the molecular testing of DTC is becoming mandatory to optimize diagnostic and therapeutical strategies. Even if the most frequent oncogenic events, occurring in a mutually exclusive manner, are BRAF V600E, followed by RAS, gene fusions mainly occurring in RET, NTRK1-3, and ALK genes have been also found. In this scenario, the definition of a molecular testing algorithm to be implemented for clinical practice is needed, in order to optimize resources and time for better clinical patient management. Methods: Gene mutations and gene fusions were evaluated on DNA and RNA extracted from cytologic or histologic samples. Mutations were tested using a lab-developed multi-gene panel able to analyze the hot-spot regions of 28 genes (total of 343 amplicons, 21.77 kb); gene fusions were tested using the Oncomine Focus assay panel. Results: Molecular characterization was performed on a total of 63 DTC (50 papillary thyroid carcinoma - PTC, 8 – follicular thyroid carcinoma - FTC, 5 – hurthle cell carcinoma - HCC) by the treating clinician for diagnostic, prognostic or clinical reasons. In 27 cases (42.8%) at least one DNA mutation was detected, 17 cases (26.9%) were positive for gene fusion rearrangement, in only 2 cases (3.1%) both DNA mutations and gene rearrangements were found and in the remaining 17 (26.9%) cases no DNA mutations or rearrangements were detected. Among mutated-subgroup, BRAF V600E mutation was the most common (14 of 63 cases – 22.2%), followed by RAS mut (9 cases – 14.2%: 4 NRAS, 3 HRAS, and 3 KRAS), both as single event in 7 and 4 cases respectively. Substitutions in TERT promoter region was found in 16 cases (25.3%), of which 14 as secondary event. Other mutations in TP53 (4 cases –6.3 %, of which 2 as single event), PIK3CA (1 case – 1.5%), AKT1(1 case – 1.5%), and RNF43 (1 case – 1.5%) were detected. Among riarranged subgroup, RET fusions and NTRK1-3 was found in 8 cases respectively (12.6%), while ALK fusion was found 1 case (1.5%). The concomitant mutational and fusion events found in 2 cases were the following: 1) HRAS and PAX8/PPARG; 2) PIK3CA/TP53/TERT and RET/CCDC6. Conclusions: As expected, BRAF and RAS mutations represent the most common genetic events in DTC and occur in a mutual exclusive manner. However, in about 30% of patients, mostly of young age, with papillary histology and lymph nodal involvement, a clinically actionable gene rearrangement has been found. Thus, we propose a two-step molecular testing algorithm with a first-level test to identify the most common mutations and a second-level test, including less common mutations and gene rearrangements, if the genes analyzed with the first level test are wild-type. The second-level test may be performed up-front in cases where there are clinicopathologic features suggestive of fusion gene TC gene or in clinically aggressive unconventional cases for which molecular therapy is being considered.